Cell survival and cell death of lymphocytes were found to be intimately connected with new vitamin A metabolites: Two naturally occurring metabolites, 14-hydroxy-retro-retinol (14HRR) and 13,14-dihydroxy-retinol (DHR), are critical for survival of lymphocytes and others cells. A third metabolite, anhydroretinol (AR) induces rapid programmed cell death. The overall goal of this renewal application is to decipher the pathway leading to the critical checkpoint where cell survival/death decisions are made. In lymphocytes, 14HRR and DHR appear to function as second messengers in signal transduction. By use of a retinoid-affinity matrix the serine/threonine kinases c-Raf-1 and several PKC isoforms have been identified as likely receptors. In recombinant proteins the N-terminal domain of c-Raf-1 and PKC contain high-affinity retinoid binding sites. The have been mapped to the cysteine-rich "zinc-finger" domains, shared by several serine/threonine kinases. Moreover, in intact cells the function of c-Raf-1 has been linked to retinoid status, and therefore c-Raf-1 qualifies as a retinoid receptor. The overall goal of the application is to verify the hypothesis that 14HRR and DHR act as natural regulators of c-Raf-1 function, promoting cell survival, and that AR, as antagonist, disrupts this regulation and incites apoptosis. As the crucial site of action must reside at a step between initial Raf-1 activation and the mitochondrial survival/death checkpoint it is proposed to investigate: in Aim 1 the Raf phosphorylation pattern, translocation and kinase competence; in Aim 2 further possible anchor proteins besides ras; in Aim 3 the consequences of disposition and function of Bcl family member proteins; in Aim 4 the hypothesis that retinoids increase the recognition and adhesion of Raf-1 to target proteins and thereby regulate substrate selection. Successful completion of the project would create a new paradigm how lipid second messengers work, with a major impact on signal transduction in lymphocytes, and with a realistic potential for intervention in signalling steps with designed retinoids that could have applications in inflammation and cancer.